The basic architecture of the vertebrate nervous system is divided into separate domains, the peripheral nervous system (PNS) and the central nervous system (CNS), that are connected by axons that travel through the boundary of these domains and creates a directional flow of information that controls the vertebrate body. Although the action potentials along axons that carry this information freely pass the CNS/PNS boundary, glial cells, which are essential for proper function of the axons, are not permitted to transverse this boundary. This restriction demarcates the two myelinating glial subtypes of the nervous system, with oligodendrocytes restricted to the CNS and Schwann cells limited to the PNS. How certain cell types are permitted to transverse the boundary while axons freely navigate across or the functional significance of separating these cells to specific domains is unknown. The importance of this boundary is underscored by the discovery that ectopically-located cells have been visualized in multiple neurological diseases including multiple sclerosis. In order for us to understand how this boundary is established and maintained to produce a functional neuronal circuit, we must evaluate its development in a way that allows us to visualize the dynamic interaction of the cells at the boundary. For this reason, I chose to utilize a model system that allows me to visualize cell-cell interactions before, during and after specific manipulation of single cells in an intact animal. The long-term goal of this proposal is to understand the development of the boundary between the CNS and PNS. Preliminary data from this system suggests that a previously unidentified cell-type that originates from the CNS, migrates through the CNS/PNS boundary where motor axons exit the spinal cord and occupies the PNS where it restricts CNS-located glia from exiting the spinal cord. Whether this same interaction also controls at the other CNS/PNS boundary region in the spinal cord will be further investigated by: 1. Characterizing which glial cell-types are located in the PNS and CNS at the CNS/PNS boundary that is located where PNS sensory axons travel into the spinal cord. Time-lapse imaging, and pharmacological/genetic ablation of glial cell precursors will give a detailed understanding of the origin of boundary glial cells and their cellular dynamics during development. 2. Investigating the cell-cell interactions of glial cells during boundary establishment and the consequence of their removal during this process. 3. Revealing the molecular requirement of Plexin/Semaphorin signaling for establishing and maintaining the glial boundary.